Plant lipids play a major role as structural elements in membranes, as cell signalling components and are the basis for petroleum replacement products, such as renewable biofuels. Renewable biofuels require the development of crops and plant species with high yields of appropriate feedstocks, for example those producing very long chain fatty acids (VLCFAs), and especially unsaturated VLCFAs. Alternatively some specifications may call for short chain fatty acids or fatty acids that have specific functional groups such as an epoxy group, an alcohol group or an acetylene bond, to name a few. All of these specialized fatty acids may be produced by introduction of one or more enzymes capable of modifying fatty acid metabolism and/or biosynthesis to produce the desired compounds.
There are few documented cases of using transgenic means to increase the oil content of Very Long Chain Fatty Acids (VLCFA) in plants. Increases in the proportion of some fatty acids for edible oils have been observed by the introduction of various plant fatty acid biosynthesis and acyltransferase genes in oilseeds. Some examples of such processes are reported in Voelker et al., Science, 257: 72-74, 1992; Voelker et al. The Plant Journal, 9: 229-241, 1996; Lassner et al. The Plant Cell, 8: 281-292, 1996; and Millar and Kunst. Plant J, 12: 121-131, 1997).
Knutzon et al. (Proc. Nat'l Acad. Sci. USA, 89: 2624-2628, 1992) reported increased stearic acid content in Brassicaceae by expressing an anti-sense construct to the stearoyl-ACP Δ9 desaturase. Hitz et al. (Proc. 9th International Cambridge Rapeseed Congress UK, pp. 470-472, 1995) reported increased proportions of oleic acid in B. napus by co-suppression of plant microsomal FAD2 (Δ12) desaturase. U.S. Pat. No. 5,824,858 reported increased proportions of 12:0 or 22:1 in the sn-2 position of triacylglycerols (TAGs) in rapeseed by expression of coconut or meadowfoam lyso-phosphatidic acid acyltransferases (LPATs; E.C. 2.3.1.51, respectively). Lassner et al. (The Plant Cell, 8: 281, 1996) and U.S. Pat. No. 5,445,947 reported increased levels of erucic acid in low erucic acid B. napus (canola) cultivars expressing a Jojoba “elongase” 3-keto-acyl-CoA synthase gene; however, the effect following elongase expression in high erucic acid cultivars was negligible, suggesting a metabolic limit.
The contributions of fatty acid elongase FAE1 and yeast L-Phosphatidyl Acyl transferase SLC1 on the fatty acid erucic acid have been separately assessed in transgenic plants (Katavic et al. Biochem Soc Trans, 28: 935-938, 2000; Katavic et al. Crop Science, 41: 739-747, 2001). Further, Mietkiewska et al. (Plant Physiology, 136: 2665, 2004; and U.S. Pub. No.: 20070204370) observed modified oil profiles and VLCFA composition in transgenic Brassica expressing a Nasturtium FAE1 gene.
The above reports demonstrated single gene transformations that only modified oil profiles by a few percent and no commercially relevant plant variety has resulted. Further the reports relied on plant transformation using Agrobacterium, with its inherent limitations, such as low copy number random integration of the transgene into the plant cell genome.
Modification of plant fatty acids or oils is an important goal, as many plant species, including crops, oilseed crops and algae species are considered as important sources of fatty acids for renewable fuels and feedstock sources for the manufacture of goods.